Streptococcus gordonii are pioneer colonizers of dental plaque biofilms. Their accumulation on surfaces is facilitated by glucans synthesized from dietary sucrose by the single extracellular glucosyltransferase (Gtf) enzyme. Regulation of the S. gordonii Gtf structural gene, gtfG, is controlled by the upstream gene, rgg which encodes a transcriptional activator that was the first-identified member of the Rgg family of regulatory proteins. There are seven Rgg-family proteins encoded in the S. gordonii chromosome including the partially characterized RggD which may be involved in glucan-independent biofilm formation. Genome sequence data have shown the widespread occurrence of rgg-family genes among Gram-positive bacteria; data are emerging about shared and unique features of the Rgg family. Rgg proteins usually control transcription of adjacent genes which are involved in various biological functions including colonization, virulence and responses to environmental conditions. Rgg-family proteins can increase or decrease expression of their target genes, often in a growth phase-dependent fashion. Some Rggs control not only the expression of adjacent genes, but act as global regulators acting at promoters of multiple genes located throughout the chromosome as occurs in Streptococcus pyogenes. Our data strongly suggest that S. gordonii Rgg may play similar global regulatory roles affecting expression of genes in addition to gtfG. An exciting recent finding in non-oral streptococcal species is that regulation by some Rgg proteins is mediated by peptide signals that function in a pheromone-like manner and act as quorum sensing molecules to coordinate the bacterial community response. These pheromones are processed from pre-peptides that are often encoded by open reading frames located near rgg genes; these peptides can positively or negatively affect the activity of the cognate Rgg. This newly-discovered mechanism of gene regulation holds intriguing possibilities for exploring novel mechanisms of oral biofilm control. Based upon the emerging evidence for pheromone interactions with Rgg proteins, we hypothesize that Rgg-family regulators may share common mechanisms of action and that as-yet-unidentified peptides may influence the activity of the S. gordonii Rgg proteins. The proposed studies will examine this possibility and provide additional insights into the function of S. gordonii Rgg family proteins via the following Specific Aims: 1) t identify and perform a preliminary characterization of selected S. gordonii target genes controlled by rgg-family determinants by comparing gene expression and resulting phenotypes in parental and mutant strains; 2) to determine if activities of S. gordonii Rgg proteins are influenced by potential pheromone-like peptides by examination of Rgg activity in the presence of both culture filtrates and synthetic peptides. Data will be used as a foundation for a future R01 application. It is hoped that these studies will identify novel facets of oral biofilm formatio that may also provide generally- applicable insights into genetic regulation by Rgg-family proteins in other Gram-positive bacteria.